ABSTRACT
The mammalian dentition is a serially homogeneous structure that exhibits wide numerical and morphological variation among multiple different species. Patterning of the dentition is achieved through complex reiterative molecular signaling interactions that occur throughout the process of odontogenesis. The secreted signaling molecule Sonic hedgehog (Shh) plays a key role in this process, and the Shh coreceptor growth arrest-specific 1 (Gas1) is expressed in odontogenic mesenchyme and epithelium during multiple stages of tooth development. We show that mice engineered with Gas1 loss-of-function mutation have variation in number, morphology, and size of teeth within their molar dentition. Specifically, supernumerary teeth with variable morphology are present mesial to the first molar with high penetrance, while molar teeth are characterized by the presence of both additional and absent cusps, combined with reduced dimensions and exacerbated by the presence of a supernumerary tooth. We demonstrate that the supernumerary tooth in Gas1 mutant mice arises through proliferation and survival of vestigial tooth germs and that Gas1 function in cranial neural crest cells is essential for the regulation of tooth number, acting to restrict Wnt and downstream FGF signaling in odontogenic epithelium through facilitation of Shh signal transduction. Moreover, regulation of tooth number is independent of the additional Hedgehog coreceptors Cdon and Boc, which are also expressed in multiple regions of the developing tooth germ. Interestingly, further reduction of Hedgehog pathway activity in Shhtm6Amc hypomorphic mice leads to fusion of the molar field and reduced prevalence of supernumerary teeth in a Gas1 mutant background. Finally, we demonstrate defective coronal morphology and reduced coronal dimensions in the molar dentition of human subjects identified with pathogenic mutations in GAS1 and SHH/GAS1, suggesting that regulation of Hedgehog signaling through GAS1 is also essential for normal patterning of the human dentition.
Subject(s)
Hedgehog Proteins , Tooth, Supernumerary , Animals , Cell Cycle Proteins/genetics , Cell Cycle Proteins/metabolism , Dentition , GPI-Linked Proteins , Gene Expression Regulation, Developmental , Hedgehog Proteins/metabolism , Humans , Mammals/genetics , Mammals/metabolism , Mice , Odontogenesis , Signal Transduction/physiology , Tooth, Supernumerary/geneticsABSTRACT
We have previously shown that PKD1, the gene encoding Polycystin-1 (or TRPP1) is expressed in human odontoblasts and that this protein is localized at the primary cilium of the cell. Nevertheless, its function remain unclear in this cell even if studies on osteoblasts, osteocytes and chondrocytes give TRPP1 as a promising candidate for mechanotransduction in response to mechanical stress. Consequently, to evaluate the role of TRPP1 in this transduction process, we needed first to generate an in vitro murine model down expressing Pkd1. Using lentivirus-mediated shRNA technology, we obtained a 60% suppression of Pkd1 mRNA expression in transfected MO6-G3 cells associated with a decrease of cell proliferation. Thus, establishment of this murine odontoblast model underexpressing Pkd1 associated with applied mechanical forces (compression or shear stress) will allow us to go further in the determination of TRPP1 involvement in odontoblasts mechanotransduction.
Subject(s)
Gene Expression Regulation/genetics , Gene Silencing/physiology , Odontoblasts/metabolism , RNA, Small Interfering/genetics , TRPP Cation Channels/genetics , Animals , Cell Culture Techniques , Cell Proliferation , Cells, Cultured , Cilia/metabolism , Gene Knockdown Techniques , Genetic Vectors/genetics , Lentivirus/genetics , Mechanotransduction, Cellular/genetics , Mechanotransduction, Cellular/physiology , Mice , Models, Animal , TransfectionABSTRACT
A primary cilium, a sensory organelle present in almost every vertebrate cell, is regularly described in odontoblasts, projecting from the surfaces of the cells. Based on the hypothesis that the primary cilium is crucial both for dentin formation and possibly in tooth pain transmission, we have investigated the expression and localization of the main cilium components and involvement of the OFD1 gene in tooth morphogenesis. Odontoblasts in vitro express tubulin, inversin, rootletin, OFD1, BBS4, BBS6, ALMS1, KIF3A, PC1, and PC2. In vivo, cilia are aligned parallel to the dentin walls, with the top part oriented toward the pulp core. Close relationships between cilium and nerve fibers are evidenced. Calcium channels are concentrated in the vicinity of the basal body. Analysis of these data suggests a putative role of cilia in sensing the microenvironment, probably related to dentin secretion. This hypothesis is enhanced by the huge defects observed on molars from Ofd1 knockout mice, showing undifferentiated dentin-forming cells.
